[Histonet] RE: Melanin problem

From:

Gayle Haider

Message: 6
Date: Fri, 27 Oct 2006 09:39:44 -0500
Hi Mel,
You might want to try an azure B counterstain if you are staining the target
antigen with DAB. Azure blue will stain mealnin a blue-green, leaving your
target antigens brown from the DAB.
Gayle Haider, CLS, MT(ASCP)
Technical Support Group
Dako North America, Inc.
6392 Via Real
Carpinteria, CA 93013
Phone: 805/566-6655 ext. 5323
From: MVaughan4@ucok.edu
Subject: [Histonet] Melanin problem
To: histonet@lists.utsouthwestern.edu
Message-ID:
Content-Type: text/plain; charset="US-ASCII"
I don't recall this question, but I would be interested in how you get
around the problem of epidermal pigmentation if you are looking for protein
expression in the epidermis; do you bleach the tissue (and if so, does that
affect antigenicity of the proteins), or can you choose a detection
chromogen that has a wild color, like a Barney purple color?
Thanks for your help.
Mel
Melville B. Vaughan, Ph. D.
Assistant Professor
Department of Biology
University of Central Oklahoma
100 N. University Drive
Edmond, OK 73034
http://www.biology.ucok.edu/PersonalPages/mvaughan/Default.htm
Gayle Haider, CLS, MT(ASCP)
Technical Support Group
Dako North America, Inc.
6392 Via Real
Carpinteria, CA 93013
Phone: 805/566-6655 ext. 5323
-----Original Message-----
From: histonet-bounces@lists.utsouthwestern.edu
[mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of
histonet-request@lists.utsouthwestern.edu
Sent: Friday, October 27, 2006 10:13 AM
To: histonet@lists.utsouthwestern.edu
Subject: Histonet Digest, Vol 35, Issue 53
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Today's Topics:
1. RE: Steamed About Steamer
(Marshall Terry Dr, Consultant Histopathologist)
2. Re: talking of unsolicited emails....... (Geoff McAuliffe)
3. Fwd: In Argentina, Pathologists Don???t Operate Clinical
Laboratories (Rene J Buesa)
4. 88331 and 88332 (Demarinis, Carolyn)
5. Antibody dilutions and temperature (MVaughan4@ucok.edu)
6. Melanin problem (MVaughan4@ucok.edu)
7. Steamer (Eric Sulkosky)
8. RE: Melanin problem (Liz Chlipala)
9. RE: 88331 and 88332 (Zajic Kari)
10. Re: Antibody dilutions and temperature (Rene J Buesa)
11. Re: Melanin problem (Rene J Buesa)
----------------------------------------------------------------------
Message: 1
Date: Fri, 27 Oct 2006 14:23:33 +0100
From: "Marshall Terry Dr, Consultant Histopathologist"
Subject: RE: [Histonet] Steamed About Steamer
To:
Message-ID:
<63984BC3A63FF542AF6EF0A237F38F4D7E4C61@LIL.xRothGen.nhs.uk>
Content-Type: text/plain; charset="iso-8859-1"
By one of those odd coincidences, the next e-mail I received after this one
was from my wife who is house sitting whilst her son and family are away,
because they have the builders in.
It included inter alia:
"The rain stopped at midday and it is a lovely afternoon. The roofers are
steaming nicely!"
Shall I ask her to find out the temperature of boiling water on the roof,
and how long they took to get to "steaming"?
Have a good weekend everyone.
PS Wonder if there are any rice cooker firms listening - I feel a pit>hole
coming on.
Dr Terry L Marshall, B.A.(Law), M.B.,Ch.B.,F.R.C.Path Consultant
Pathologist Rotherham General Hospital South Yorkshire England
terry.marshall@rothgen.nhs.uk
-----Original Message-----
From: Fred Underwood [mailto:funderwood@mcohio.org]
Sent: 27 October 2006 13:22
To: amosbrooks@gmail.com; histonet@lists.utsouthwestern.edu; Marshall Terry
Dr, Consultant Histopathologist
Subject: RE: [Histonet] Steamed About Steamer
I'm happy to see that this has become a "Nan" issue.
>>> "Marshall Terry Dr,Consultant Histopathologist"
10/27/2006 5:40 AM >>> I am intrigued by
this discussion.
I have been using rice steamers for 42 years.
I use them to cook rice, oddly enough:-) They have been used with great
frequency - I love my Indian food.
Never ever, have I had one that decreased in performance.
The only problem has been that the inner pan gets gradually pitted by the
salt until it holes.
Dr Terry L Marshall, B.A.(Law), M.B.,Ch.B.,F.R.C.Path Consultant
Pathologist Rotherham General Hospital South Yorkshire England
terry.marshall@rothgen.nhs.uk
-----Original Message-----
From: Amos Brooks [mailto:amosbrooks@gmail.com]
Sent: 27 October 2006 01:06
To: histonet@lists.utsouthwestern.edu
Subject: [Histonet] Steamed About Steamer
Hi,
I have only used 2 brands of steamer so far but there's no reason
another can't work as well. Certainly steamers can wear out as one user has
pointed out. I have seen this, the heating element stops getting the
temperature high enough. As this is a new steamer that shouldn't be the
problem.
It's possible that the steaming rack is too far from the element
leaving the steam enough time to cooldown before it reaches the slides.
That
is pretty much a design flaw if that is the case. If so there's nothing else
for it but to replace it. Try to get something compact that allows enough
space for whatever container you are using to hold the slide rack.
It is possible that the vents are allowing the steam out too quickly.
That could prevent the steam from heating the buffer up enough. If so you
could cover some of the holes (Try duct tape at first, you can figure out
something more permanent if it works.
To make sure the buffer gets up to the right temperature, I have holes
drilled in the top of my steamers just big enough to fit a thermometer.
This
way I know the buffer is hot enough to put the slides in and when the buffer
gets back up to the right temperature after the slides are added. This is
great because you can directly monitor the temperature of the solution in
real time thruought the retrieval process. (Try that in a microwave or
pressure cooker)
In response to the gentleman that was having trouble with E-Cadherin,
we have found an extra 10 min. produces a stronger antibody reaction.
This
works for ER & PR too. I am sure we'll find more if we actually get time to
test more antibodies.
Good Luck
Amos Brooks
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------------------------------
Message: 2
Date: Fri, 27 Oct 2006 09:39:15 -0400
From: Geoff McAuliffe
Subject: Re: [Histonet] talking of unsolicited emails.......
To: "Andrea T. Hooper"
Cc: Histonet
Message-ID: <45420C03.9010607@umdnj.edu>
Content-Type: text/plain; format=flowed; charset=ISO-8859-1
I got it too and I also avoid dealing with spammers.
Geoff
Andrea T. Hooper wrote:
> Indeed I did and it irritated me. I make a conscious effort not to
> purchase from companies who solicit business by stealing information
> off of a listserv and spamming email accounts .... so quite honestly
> it's their loss.
>
>
>> Anyone else get the email from Patrick Levasseur, Econo-Lab
>> [info@econo-lab.com]? They have clearly harvested emails from
>> histonet as I got the mail to my home and work emails, both of which
>> are subscribed to histonet.
>>
>> I am more than happy for vendors to be on the list as they often have
>> solutions to things and offer help but I do think harvesting emails
>> and putting them on a mailing list for their company is out of order.
>> Apparently
>> I have to email to be taken off their mailing list too!
>>
>> Alan Bishop
>> Charge scientist
>> Histology
>> Medlab Central
>> Palmerston North
>> New Zealand
>>
>> Tel: 06 952 3135
>> Fax: 06 952 3199
>>
>
--
--
**********************************************
Geoff McAuliffe, Ph.D.
Neuroscience and Cell Biology
Robert Wood Johnson Medical School
675 Hoes Lane, Piscataway, NJ 08854
voice: (732)-235-4583; fax: -4029
mcauliff@umdnj.edu
**********************************************
------------------------------
Message: 3
Date: Fri, 27 Oct 2006 06:49:28 -0700 (PDT)
From: Rene J Buesa
Subject: [Histonet] Fwd: In Argentina, Pathologists Don???t Operate
Clinical Laboratories
To: histonet@lists.utsouthwestern.edu
Message-ID: <20061027134928.79568.qmail@web61215.mail.yahoo.com>
Content-Type: text/plain; charset=iso-8859-1
Dear Colleagues:
I just received this very interesting report that I want to share will
all.
René J.
DARK DAILY wrote:
From: "DARK DAILY"
To: rjbuesa@yahoo.com
Subject: In Argentina, Pathologists Donâ€™t Operate Clinical Laboratories
Date: Fri, 27 Oct 2006 13:30:30 +0000
If you are having trouble viewing this email or any of the pages
that follow, please open a new browser window and visit:
http://www.darkdaily.com/daily/daily_email10.27.06.htm
Add the Dark Daily to your address book and never miss an email!
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Home | Archive | About | Join--
In Argentina, Pathologists Don't Operate Clinical
Laboratories And Patients Deliver Their Own Lab Test Reports to Their
Doctors
DATELINE—BUENOS AIRES, ARGENTINA: Believe it or not, Argentina is a
country where pathologists are not associated with clinical laboratory
testing! I am Argentina this week to speak to a group of laboratory owners,
speak to health insurance executives, and visit clinical laboratories. My
hosts on this trip are Sysmex Corporation and Roche Diagnostics Argentina.
The most startling thing that I learned is that pathologists do not own
clinical laboratories in Argentina, nor do they manage clinical laboratories
in hospitals. That responsibility is handled by clinical biochemists.
Further, interaction between anatomic pathologists and clinical laboratories
is relatively limited. This is a much different situation than the common
clinical models and business arrangements used in the United States, Canada,
and similar nations.
Another interesting difference is that doctors give their patients the
laboratory test requisition. Patients then show up at the laboratory of
their choice, where specimens are collected. When the results are ready, the
laboratory contacts the patient, who comes by the laboratory or patient
service center, picks up a paper copy of his or her test report, and then
personally delivers the lab test report to his or her physician. This
arrangement is counterintuitive to the American system. On the other hand,
it does provide a powerful example that patients can reliably play a role
this aspect of their healthcare.
Competition is intense among private laboratories in Buenos Aires,
Argentina. There are private health insurance companies. Labs that bid for
provider status in these contracts tend to keep the prices for lab testing
at unprofitably low levels. Another factor in the financial struggles of the
entire health system in Argentina was the devaluation of the Argentinian
Peso in 2001. The purchasing power of the Peso fell by 66% overnight! This
has made the purchase of laboratory testing instruments, reagents, and other
laboratory consumables prohibitively expensive for labs in Argentina. The
effects of this devaluation are still dragging down the finances of this
countries' lab industry.
During this week, I gained many useful insights about laboratory management,
different business models for laboratory testing, and the opportunities for
laboratories in the United States to benefit from some of the experience
provided by the laboratory testing marketplace in Argentina. These are
examples of how innovations by laboratories in one country can be adopted by
labs in other countries. I hope to share some of the more powerful
innovations with you in coming weeks.
Reporting from Buenos Aires, your faithful Editor, Robert L. Michel
(E-mail Robert at rmichel@darkdaily.com or schristensen@darkdaily.com.)
---------------------------------
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Message: 4
Date: Fri, 27 Oct 2006 10:14:06 -0400
From: "Demarinis, Carolyn"
Subject: [Histonet] 88331 and 88332
To:
Message-ID:
Content-Type: text/plain; charset="US-ASCII"
There seems to be some confusion on what codes to use when multiple frozens
are sent on multiple specimens.
I need some clarification, therefore, I am providing an example.
If two specimens are sent to the lab for frozen section labeled:
A) left upper eyelid 12 o'clock marker
B) left upper eyelid
Frozen section diagnosis on A) is basal cell carcinoma at margins.
Frozen section diagnosis on B) is no evident malignancy.
Am I correct in using 88331 x 2 in addition to 88305 x 2?
I am getting billing rejections stating that I should be using 88331 x 1,
88332 x 1, 88305 x 2.
If so, please provide me with appropriate documentation.
Thank you.
------------------------------
Message: 5
Date: Fri, 27 Oct 2006 09:33:36 -0500
From: MVaughan4@ucok.edu
Subject: [Histonet] Antibody dilutions and temperature
To: histonet@lists.utsouthwestern.edu
Message-ID:
Content-Type: text/plain; charset="US-ASCII"
Hello all,
I have noticed there are different diluents used for various antibodies; I
have seen PBS, PBS plus albumin, and PBS/albumin/Tween.
What is the difference between using one or the other of these? Also, how
does incubation temperature play a role in the choice? Thanks for your
answer, Tim :). Actually I would like to hear anyone's opinion since just
about everyone here is an IHC or IF expert. Enjoy the weekend!
Mel
Melville B. Vaughan, Ph. D.
Assistant Professor
Department of Biology
University of Central Oklahoma
100 N. University Drive
Edmond, OK 73034
http://www.biology.ucok.edu/PersonalPages/mvaughan/Default.htm
------------------------------
Message: 6
Date: Fri, 27 Oct 2006 09:39:44 -0500
From: MVaughan4@ucok.edu
Subject: [Histonet] Melanin problem
To: histonet@lists.utsouthwestern.edu
Message-ID:
Content-Type: text/plain; charset="US-ASCII"
I don't recall this question, but I would be interested in how you get
around the problem of epidermal pigmentation if you are looking for protein
expression in the epidermis; do you bleach the tissue (and if so, does that
affect antigenicity of the proteins), or can you choose a detection
chromogen that has a wild color, like a Barney purple color?
Thanks for your help.
Mel
Melville B. Vaughan, Ph. D.
Assistant Professor
Department of Biology
University of Central Oklahoma
100 N. University Drive
Edmond, OK 73034
http://www.biology.ucok.edu/PersonalPages/mvaughan/Default.htm
------------------------------
Message: 7
Date: Fri, 27 Oct 2006 10:54:41 -0400
From: "Eric Sulkosky"
Subject: [Histonet] Steamer
To:
Message-ID: <57A8B4592C634A4CAD3A6CB1A0F8103B34DED9@mhsmail2.mhs.org>
Content-Type: text/plain; charset="us-ascii"
Hi All
My laboratory has been using the rice steamer for years and has not had any
problems. I would like to ask, have you been preheating your solutions
before trying to retrieve? You mentioned that you were having problems with
TTF and Ecad. Are these Dako antibodies? We have found the best results with
retrieving, when you preheat your solutions for twenty minutes. Also what pH
of solutions are you using?
Our procedure for retrieving is as follows:
1. Decide which pH needs to be used. In our laboratory we use High
pH 9.9 and Regular pH 6.0
2. For the antibodies you mentioned Ecad and TTF we use the High
pH. We have much better results.
3. Set up your steamer, pour your Target Retrieval and preheat in
steamer for twenty minutes.
4. For High pH put your slides in the preheated solution and set
your timer for 40minutes. Remove the slides from the steamer and leave in
your Target Retrieval. Set another clock for 20minutes and let the solution
come to room temperature. After it is room temperature, transfer the slides
into buffer and let them sit for 5 minutes. This helps from your tissue
drying out when staining.
5. For Regular pH put your slides in the preheated solution and set
your timer for 20minutes. Remove the slides from the steamer and leave in
your Target Retrieval. Set another clock for 20minutes and let the solution
come to room temperature. After it is room temperature, transfer the slides
into buffer and let them sit for 5 minutes. This helps from your tissue
drying out when staining.
6. Then proceed with your staining.
Important note: It is very important to let your slides sit in the retrieval
and come to room temperature before transferring them into buffer or water.
The cooling down is built into the retrieval process, so I would advise not
to directly transfer into the buffer right out of the hot steamer.
I hope this helps.
Eric
sulkoskye@monhealthsys.org
------------------------------
Message: 8
Date: Fri, 27 Oct 2006 09:12:12 -0600
From: "Liz Chlipala"
Subject: RE: [Histonet] Melanin problem
To: ,
Message-ID: <002101c6f9da$477f9c50$0300a8c0@domain.Premier>
Content-Type: text/plain; charset="us-ascii"
Mel
When I'm dealing with endogenous brown pigment I switch to a AP detection
system and use Fast Red or another red chromagen or if you want to stay with
a HRP system then use AEC.
Liz
Elizabeth A. Chlipala, BS, HTL(ASCP)QIHC Manager Premier Laboratory, LLC
P.O. Box 18592 Boulder, CO 80308 phone (303) 735-5001 fax (303) 735-3540
liz@premierlab.com www.premierlab.com
Ship to Address:
Premier Laboratory, LLC
University of Colorado at Boulder
MCDB, Room A3B40
Boulder, CO 80309
-----Original Message-----
From: histonet-bounces@lists.utsouthwestern.edu
[mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of
MVaughan4@ucok.edu
Sent: Friday, October 27, 2006 8:40 AM
To: histonet@lists.utsouthwestern.edu
Subject: [Histonet] Melanin problem
I don't recall this question, but I would be interested in how you get
around the problem of epidermal pigmentation if you are looking for protein
expression in the epidermis; do you bleach the tissue (and if so, does that
affect antigenicity of the proteins), or can you choose a detection
chromogen that has a wild color, like a Barney purple color?
Thanks for your help.
Mel
Melville B. Vaughan, Ph. D.
Assistant Professor
Department of Biology
University of Central Oklahoma
100 N. University Drive
Edmond, OK 73034
http://www.biology.ucok.edu/PersonalPages/mvaughan/Default.htm
_______________________________________________
Histonet mailing list
Histonet@lists.utsouthwestern.edu
http://lists.utsouthwestern.edu/mailman/listinfo/histonet
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------------------------------
Message: 9
Date: Fri, 27 Oct 2006 11:09:49 -0400
From: "Zajic Kari"
Subject: RE: [Histonet] 88331 and 88332
To: "Demarinis, Carolyn" ,
Message-ID:
<095327C7CDBDF64B9E9728A54799091E015CC7A8@ORLEV03.hca.corpad.net>
Content-Type: text/plain; charset="iso-8859-1"
In my opinion (and experience) you can bill an 88331 x2 because you have two
separate specimens in two separate containers. Every additional frozen done
on ONE specimen gets charged an 88332, for example if you had done margins
on your first (A) specimen x3 frozen blocks, you can charge 88331 x1 and
88332 x2.
I am not a professional biller, but this is my understanding. I checked my
American Medical Association Surgical Pathology billing guidelines and they
(AMA) define pathology specimens as below:
"A specimen is defined as tissue or tissues that is (are) submitted for
individual and separate attention, requiring individual examination and
pathologic diagnosis. Two or more such specimens from the same patient (eg,
separately identified endoscopic biopsies, skin lesions, etc.) are each
appropriately assigned an individual code reflective of its proper level of
service."
hope this helps!!! :)
Kari Marie Zajic HTL,MLT
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-----Original Message-----
From: histonet-bounces@lists.utsouthwestern.edu
[mailto:histonet-bounces@lists.utsouthwestern.edu]On Behalf Of Demarinis,
Carolyn
Sent: Friday, October 27, 2006 10:14 AM
To: HISTONET@PATHOLOGY.SWMED.EDU
Subject: [Histonet] 88331 and 88332
There seems to be some confusion on what codes to use when multiple frozens
are sent on multiple specimens.
I need some clarification, therefore, I am providing an example.
If two specimens are sent to the lab for frozen section labeled:
A) left upper eyelid 12 o'clock marker
B) left upper eyelid
Frozen section diagnosis on A) is basal cell carcinoma at margins.
Frozen section diagnosis on B) is no evident malignancy.
Am I correct in using 88331 x 2 in addition to 88305 x 2?
I am getting billing rejections stating that I should be using 88331 x 1,
88332 x 1, 88305 x 2.
If so, please provide me with appropriate documentation.
Thank you.
_______________________________________________
Histonet mailing list
Histonet@lists.utsouthwestern.edu
http://lists.utsouthwestern.edu/mailman/listinfo/histonet
------------------------------
Message: 10
Date: Fri, 27 Oct 2006 08:10:26 -0700 (PDT)
From: Rene J Buesa
Subject: Re: [Histonet] Antibody dilutions and temperature
To: MVaughan4@ucok.edu, histonet@lists.utsouthwestern.edu
Message-ID: <20061027151026.57585.qmail@web61218.mail.yahoo.com>
Content-Type: text/plain; charset=iso-8859-1
Mel:
1- Abs + PBS: this is the neutral buffered environment the Ab should be
diluted;
2- Abs + PBS + albumen: this is the same environment but the addition of
albumen will preserve the Ab better (for a longer time);
3- Abs + PBS + albumen + Tween: the Tween, as a surfactant, will allow the
Ab to "spread" better over the section and produce a better staining.
You should dilute your Abs in PBS with aldumen (1%) + Tween 20 (0.05%) to
obtain better results.
Some instruments increase the incubation temperature to obtain quicker
results, or the incubation can be done overnight at lower temperatures to
allow for weaker dilutions and prevent evaporation. Temperature affects
incubation spead according with Van't Hoff's laws (direct effect).
When you have many experts in the same field with different opinions,
either just 1 is right, or several have a "fraction of the truth" or all are
wrong; make your pick!
René J.
MVaughan4@ucok.edu wrote:
Hello all,
I have noticed there are different diluents used for various antibodies; I
have seen PBS, PBS plus albumin, and PBS/albumin/Tween.
What is the difference between using one or the other of these? Also, how
does incubation temperature play a role in the choice? Thanks for your
answer, Tim :). Actually I would like to hear anyone's opinion since just
about everyone here is an IHC or IF expert. Enjoy the weekend!
Mel
Melville B. Vaughan, Ph. D.
Assistant Professor
Department of Biology
University of Central Oklahoma
100 N. University Drive
Edmond, OK 73034
http://www.biology.ucok.edu/PersonalPages/mvaughan/Default.htm
_______________________________________________
Histonet mailing list
Histonet@lists.utsouthwestern.edu
http://lists.utsouthwestern.edu/mailman/listinfo/histonet
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------------------------------
Message: 11
Date: Fri, 27 Oct 2006 08:14:53 -0700 (PDT)
From: Rene J Buesa
Subject: Re: [Histonet] Melanin problem
To: MVaughan4@ucok.edu, histonet@lists.utsouthwestern.edu
Message-ID: <20061027151453.86842.qmail@web61213.mail.yahoo.com>
Content-Type: text/plain; charset=iso-8859-1
I used to bleach without affecting antigenicity.
René J.
MVaughan4@ucok.edu wrote:
I don't recall this question, but I would be interested in how you get
around the problem of epidermal pigmentation if you are looking for protein
expression in the epidermis; do you bleach the tissue (and if so, does that
affect antigenicity of the proteins), or can you choose a detection
chromogen that has a wild color, like a Barney purple color?
Thanks for your help.
Mel
Melville B. Vaughan, Ph. D.
Assistant Professor
Department of Biology
University of Central Oklahoma
100 N. University Drive
Edmond, OK 73034
http://www.biology.ucok.edu/PersonalPages/mvaughan/Default.htm
_______________________________________________
Histonet mailing list
Histonet@lists.utsouthwestern.edu
http://lists.utsouthwestern.edu/mailman/listinfo/histonet
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Get on board. You're invited to try the new Yahoo! Mail.
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End of Histonet Digest, Vol 35, Issue 53
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